Purpose: Nerve grafting has been the most widely accepted surgical intervention for the repair of peripheral nerve defects. Traditionally however, such interventions have been associated with varying degrees of donor site morbidity. In the search for alternative methods of peripheral nerve repair we have experimented with the use of biological and artificial materials for the creation of nerve conduits. This study was performed to assess the effects of bone marrow stromal cells (BMSC) in nerve gaps repaired with isogenic epineural tubes filled with isogenic and allogenic BMSCs.
Methods: A total of 54 epineural tubes were transplanted in 3 experimental groups (18 animals in each group). Group 1 was control saline, Group 2 isogenic BMSCs and Group 3 allogenic BMSCs. In Groups 2 and 3, BMSCs were stained with PKH-dye before transplantation, to assess nerve engraftment and migration. After staining 2.5-3.0 x 106 cells were delivered directly into the transplanted epineural tube. Evaluations were performed at 6, 12 and 18 weeks post-transplant. Sensory and motor recoveries were evaluated by Gastrocnemius Muscle Index (GMI) ,pinprick, toe-spread and Somato-Sensory Evoked Potentials (SSEP). Axonal countings were performed in addition to immunostaining with nerve growth factors: NGF, Laminin B2, GFAP, VEGF and Von Willebrand Factor for the assessment of the expression of neurotrophic factors and regenerative potential of transplanted BMSCs.
Results: 6 weeks post transplantation all groups scored 3 on the pin-prick test. Toe spread for groups 1,2 and 3 were respectively 1.7; 2; 1. SSEPs in groups 1, 2 and 3 (P1, N2-latencies; P1, N2 % of normal values) were respectively (20.2; 23.6; 113; 95), (17.5; 18.1; 98; 73) and (15.7; 21.65; 88; 87). GMIs in groups 1,2 and 3 were respectively (0.45; 0.48; 0.47). Group 2 showed a higher number of regenerated axons (90.6 ± 26.9) compared to Group 1 (71.4 ± 3.0) and 3 (76.4 ± 5.4). In groups 2 and 3 (with BMSCs) PKH positive cells in addition to NGF, Laminin B2, GFAP, VEGF and Von Willebrand Factor were found in the transplanted tubes. 18 weeks post transplant isogenic stromal cells (Group 2) expressed neurotrophic factors in the distal portion of the transplanted nerve in contrast to the allogenic group (Group 3) in which neurotrophic factors were expressed in the middle portion of the transplanted nerve. NGF-staining in combination with PKH-staining confirmed that BMSCs differentiated into neural tissues. Upregulation of Laminin-B2 in both groups indicated active nerve regeneration.
Conclusion: Co-transplantation of BMSCs within epineural tubes enhanced nerve regeneration of peripheral nerve defects and confirmed the regenerative potential of BMSCs through their differentiation into neural tissue. Isogenic BMSCs showed a greater regeneration potential than allogenic BMSCs. These findings correlated with functional recovery of the nerves as measured by SSEP and axonal regeneration utilizing electron microscopy.